The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9) genome editing system has revolutionized the field of genome editing for therapeutic ...applications for genetic diseases and cancers, the development of disease models, identification of targets for drug development. However, safe and efficient delivery of the CRISPR/Cas9 genome editing system in vivo remains a significant challenge. Compared to viral vectors, non-viral nano-vectors have the following advantages: cost-effective, scale-up production, ease of chemical modification, large packaging capacity, lower immunogenicity, better protection of CRISPR/Cas9 genome edition system from degradation in vivo. This comprehensive Review article highlights the recent advances in delivering the three categories of CRISPR/Cas9 genome editing cargoes (i.e., Cas9 DNA plasmid, Cas9 mRNA, and Cas9 ribonucleoprotein) by using three types of nano-vectors: lipid-based, polymer-based, and polymer inorganic nano-vectors to treat cancers and genetic diseases via two major gene-editing pathways: non-homologous end joining pathway and homology-directed repair pathway. This article also notes the therapeutic applications for treating cancers and genetic diseases, provides insights into the challenges of nano-vectors for the delivery of the CRISPR/Cas9 genome editing system and proposes promising strategies in future development, especially in the final clinical translation.
We report recent advances in non-viral nano-vector delivery of the CRISPR/Cas9 genome editing payloads (i.e., Cas9 DNA plasmid, Cas9 mRNA, and Cas9 RNP), showing as potential therapeutic applications for treating cancers and genetic diseases via systemic or local administration into organs, such as brain, eye, lung, liver, muscle, and tumor. Display omitted
•Recent advances in non-viral nano-vector delivery of the CRISPR/Cas9 genome editing payloads is comprehensively reviewed.•Lipid, polymer, or inorganic composition are typical examples of nano-vectors.•Nano-vectors can deliver three different payloads including plasmid, mRNA, and RNP for CRISPR/Cas9 genome editing.•More research is expected to explore the next-generation nano-vectors.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Chronic white adipose tissue (WAT) inflammation has been recognized as a critical early event in the pathogenesis of obesity‐related disorders. This process is characterized by the increased ...residency of proinflammatory M1 macrophages in WAT. However, the lack of an isogenic human macrophage‐adipocyte model has limited biological studies and drug discovery efforts, highlighting the need for human stem cell‐based approaches. Here, human induced pluripotent stem cell (iPSC) derived macrophages (iMACs) and adipocytes (iADIPOs) are cocultured in a microphysiological system (MPS). iMACs migrate toward and infiltrate into the 3D iADIPOs cluster to form crown‐like structures (CLSs)‐like morphology around damaged iADIPOs, recreating classic histological features of WAT inflammation seen in obesity. Significantly more CLS‐like morphologies formed in aged and palmitic acid‐treated iMAC‐iADIPO‐MPS, showing the ability to mimic inflammatory severity. Importantly, M1 (proinflammatory) but not M2 (tissue repair) iMACs induced insulin resistance and dysregulated lipolysis in iADIPOs. Both RNAseq and cytokines analyses revealed a reciprocal proinflammatory loop in the interactions of M1 iMACs and iADIPOs. This iMAC‐iADIPO‐MPS thus successfully recreates pathological conditions of chronically inflamed human WAT, opening a door to study the dynamic inflammatory progression and identify clinically relevant therapies.
Here, iPSC‐derived macrophages and adipocytes are cocultured in a microphysiological system. Macrophages migrate toward and infiltrate into the 3D adipocytes cluster to form crown‐like structures. M1 but not M2 macrophages induced insulin resistance and dysregulated lipolysis in adipocytes, causing a reciprocal proinflammatory loop. This isogenic system opens a door to study the dynamic inflammatory progression and identify clinically relevant therapies.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Impaired white adipose tissue (WAT) function has been recognized as a critical early event in obesity‐driven disorders, but high buoyancy, fragility, and heterogeneity of primary adipocytes have ...largely prevented their use in drug discovery efforts highlighting the need for human stem cell‐based approaches. Here, human stem cells are utilized to derive metabolically functional 3D adipose tissue (iADIPO) in a microphysiological system (MPS). Surprisingly, previously reported WAT differentiation approaches create insulin resistant WAT ill‐suited for type‐2 diabetes mellitus drug discovery. Using three independent insulin sensitivity assays, i.e., glucose and fatty acid uptake and suppression of lipolysis, as the functional readouts new differentiation conditions yielding hormonally responsive iADIPO are derived. Through concomitant optimization of an iADIPO‐MPS, it is abled to obtain WAT with more unilocular and significantly larger (≈40%) lipid droplets compared to iADIPO in 2D culture, increased insulin responsiveness of glucose uptake (≈2–3 fold), fatty acid uptake (≈3–6 fold), and ≈40% suppressing of stimulated lipolysis giving a dynamic range that is competent to current in vivo and ex vivo models, allowing to identify both insulin sensitizers and desensitizers.
This study uses human stem cells to derive metabolically functional 3D adipose tissue in a microphysiological system. Through concomitant optimization, the adipose system shows mostly unilocular and significantly larger lipid droplets, increased insulin responsiveness of glucose uptake, fatty acid uptake, and suppressing of stimulated lipolysis compared to 2D culture, providing a good dynamic range to identify insulin sensitizers and desensitizers.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Transcriptomic profiling of tumor tissues introduces a large database, which has led to improvements in the ability of cancer diagnosis, treatment, and prevention. However, performing tumor ...transcriptomic profiling in the clinical setting is very challenging since the procurement of tumor tissues is inherently limited by invasive sampling procedures. Here, the feasibility of purifying hepatocellular carcinoma (HCC) circulating tumor cells (CTCs) from clinical patient samples is demonstrated with improved molecular integrity using Click Chips in conjunction with a multimarker antibody cocktail. The purified CTCs are then subjected to messenger RNA profiling by NanoString nCounter platform, targeting 64 HCC‐specific genes, which are generated from an integrated data analysis framework with eight tissue‐based prognostic gene signatures from seven publicly available HCC transcriptomic studies. After bioinformatics analysis and comparison, the HCC CTC‐derived gene signatures show high concordance with HCC tissue‐derived gene signatures from the Cancer Genome Atlas database, suggesting that HCC CTCs purified by Click Chips can enable the translation of HCC tissue molecular profiling into a noninvasive setting.
Covalent chemistry‐mediated multimarker purification of hepatocellular carcinoma circulating tumor cells (HCC CTCs) is developed to realize noninvasive detection of molecular signatures of HCC. The purification system enables improved purity and molecular integrity of the purified HCC CTCs for conducting transcriptomic profiling of a strictly selected HCC‐specific gene panel.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Particle image velocimetry (PIV) is a widely used image processing technique for mapping deformation in centrifuge models. One of the problems associated with PIV is the occurrence of incorrect or ‘ ...wild vectors’. A ‘ wild vector’ is defined as a vector that does not follow the general directional trend of the other vectors surrounding it, which if present in large numbers, can affect the accuracy of the deformation plot. This problem is particularly important in clay model, the surfaces of which often do not have inherent texture. The paper examines some possible enhancements to PIV images by fine tuning three parameters, namely type of texture material, method of texture placement and the use of high resolution colour imagery. The trials were conducted on centrifuge models of tunnels. The desired outcome is to minimize the percentage of wild vectors that is attained from the PIV analyses of the recorded images. The results show that with the use of colour model terrain, the likelihood of clay obscuration can be reduced significantly, Furthermore, by the use of an appropriate method of scatter will offer favorable image contrast for PIV based experiments involving the use of saturated clay. Finally the advantage of colour imagery is that it is able to capture the difference in color as the coloured camera is able to detect the hue, saturation and luminance of the texture used. These lead to a corresponding improvement in image contrast and hence, PIV analysis results.
•The GMCNs displayed higher food dye adsorption capacity and lower cytotoxicity.•Adsorption process was spontaneous and exothermic which fitted the Langmuir model.•The GMCNs were regenerated ...successfully and can be reuse for food dye adsorption.
Glutaraldehyde cross-linked magnetic chitosan nanoparticles (GMCNs) were prepared through cross-linking modification of magnetic chitosan nanoparticles (MCNs) using glutaraldehyde as crosslinker, that not only exhibited excellent food dyes adsorption performance, but also showed low cytotoxicity. Adsorption characteristics of GMCNs in FD&C Blue 1 and D&C Yellow 5 aqueous solutions have been studied and results indicated that the adsorption capacities were affected by initial pH values, initial dye concentrations and temperatures. Food dyes adsorption followed with thepseudo-second-orderreaction, and equilibrium experiments were well fitted the Langmuir isotherm model. Maximum adsorption capacities of GMCNs displayed at pH 3.0 and at 25°C, being up to 475.61 and 292.07mg/g, for FD&C Blue 1 and D&C Yellow 5 respectively. Thermodynamic results demonstrated that the adsorption processes were spontaneous and exothermic. Furthermore, it was found that the GMCNs can be regenerated and reused through dye desorption in alkaline solution.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Three prime repair exonuclease 1 (TREX1) is an essential exonuclease in mammalian cells, and numerous in vivo and in vitro data evidenced its participation in immunity regulation and in genotoxicity ...remediation. In these very complicated cellular functions, the molecular mechanisms by which duplex DNA substrates are processed are mostly elusive because of the lack of structure information. Here, we report multiple crystal structures of TREX1 complexed with various substrates to provide the structure basis for overhang excision and terminal unwinding of DNA duplexes. The substrates were designed to mimic the intermediate structural DNAs involved in various repair pathways. The results showed that the Leu24-Pro25-Ser26 cluster of TREX1 served to cap the nonscissile 5'-end of the DNA for precise removal of the short 3'-overhang in L- and Y-structural DNA or to wedge into the double-stranded region for further digestion along the duplex. Biochemical assays were also conducted to demonstrate that TREX1 can indeed degrade double-stranded DNA (dsDNA) to a full extent. Overall, this study provided unprecedented knowledge at the molecular level on the enzymatic substrate processing involved in prevention of immune activation and in responses to genotoxic stresses. For example, Arg128, whose mutation in TREX1 was linked to a disease state, were shown to exhibit consistent interaction patterns with the nonscissile strand in all of the structures we solved. Such structure basis is expected to play an indispensable role in elucidating the functional activities of TREX1 at the cellular level and in vivo.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Taiwan established a system of universal National Health Insurance (NHI) in March, 1995. Today, the NHI covers more than 98% of Taiwan's population and enrollees enjoy almost free access to ...healthcare with small co-payment by most clinics and hospitals. Yet while this expansion of coverage will almost inevitably have improved access to health care, however, it cannot be assumed that it will necessarily have improved the health of the population. The aim of this study was to determine whether the introduction of National Health Insurance (NHI) in Taiwan in 1995 was associated with a change in deaths from causes amenable to health care.
Identification of discontinuities in trends in mortality considered amenable to health care and all other conditions (non-amenable mortality) using joinpoint regression analysis from 1981 to 2005.
Deaths from amenable causes declined between 1981 and 1993 but slowed between 1993 and 1996. Once NHI was implemented, the decline accelerated significantly, falling at 5.83% per year between 1996 and 1999. In contrast, there was little change in non-amenable causes (0.64% per year between 1981 and 1999). The effect of NHI was highest among the young and old, and lowest among those of working age, consistent with changes in the pattern of coverage. NHI was associated with substantial reductions in deaths from circulatory disorders and, for men, infections, whilst an earlier upward trend in female cancer deaths was reversed.
NHI was associated in a reduction in deaths considered amenable to health care; particularly among those age groups least likely to have been insured previously.
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CEKLJ, DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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•A new two-step route for nano-scaled magnetic chitosan particles preparation.•Triton X-100 reversed-phase microemulsion system was used for chitosan coating.•Narrow size distribution ...of magnetic chitosan nanoparticles was achieved.•Quantitative evaluation of recoverability for the magnetic chitosan nanoparticles.
A new two-step route for the preparation of nano-scaled magnetic chitosan particles has been developed, different from reported one-step in situ preparation and two-step preparation method of reversed-phase suspension, Triton X-100 reversed-phase water-in-oil microemulsion encapsulation method was employed in coating the pre-prepared Fe3O4 nanoparticles with chitosan. The resultant magnetic chitosan particles owned a narrow size distribution ranging from 50 to 92nm. X-ray diffraction patterns (XRD) indicated that the chitosan coating procedure did not change the spinal structure of Fe3O4 magnetic nanoparticles. The results of Fourier transform infrared (FTIR) analysis and thermogravimetric analysis (TGA) demonstrated that the chitosan was coated on Fe3O4 nanoparticles and its average mass content was ∼50%. The saturated magnetization of the magnetic Fe3O4/chitosan nanoparticles reached 18.62emu/g, meanwhile, the nanoparticles showed the characteristics of superparamagnetism. The magnetic chitosan nanoparticles showed a high recoverability of 99.99% in 10min when pH exceeded 4. The results suggested that the as-prepared magnetic chitosan particles were nano-scaled with a narrow size distribution and a high recoverability.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Increasing the level of correspondence between measures of growth mindset and their related outcomes could afford more precise prediction of the relationships between growth mindset and ...social-emotional outcomes. To illustrate the value of measurement correspondence, two studies were conducted in Hong Kong. Study 1 showed that an agent-correspondent growth mindset measure (parents’ perception of the malleability of their children's personal qualities), compared to an agent-non-correspondent one (parents’ belief in the malleability of personal qualities of a generalized other), had stronger predictive relationship with children's likelihood of displaying difficult behaviors. Study 2 found that children's self-theories about the malleability of their intelligence (an intrapersonal construct) had stronger predictive relationship with academic engagement (an intrapersonal outcome) than did their perception of growth mindset norm (a normative construct). However, perceived growth mindset norm regarding personal qualities had stronger predictive relationship with peer relationship quality (an interpersonal outcome). Together these results demonstrated that when corresponding measures of growth mindset were used to predict an outcome, more reliable growth mindset effects would emerge.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
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